Age Sensitivity of NFκB Abundance and Programmed Cell Death in Erythrocytes Induced by NFκB Inhibitors

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Background/Aims: Mitoxantrone, a cytotoxic drug used for the treatment of malignancy and multiple sclerosis, is at least in part effective by triggering apoptosis. Similar to apoptosis of nucleated cells, erythrocytes may enter eryptosis, a type of suicidal cell death. Hallmarks of eryptosis are cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Signalling involved in eryptosis include Ca²⁺-entry, ceramide formation and oxidative stress. Methods: Cell volume was estimated from forward scatter, phosphatidylserine-exposure from annexin V binding, formation of reactive oxidant species (ROS) from 2′,7′-dichlorodihydrofluorescein-diacetate fluorescence, and ceramide abundance from binding of fluorescent antibodies in flow cytometry. Results: A 48 hours exposure to mitoxantrone was followed by significant decrease of forward scatter (≥ 5 μg/ml mitoxantrone) and increase of annexin-V-binding (≥ 10 μg/ml mitoxantrone), effects paralleled by significant increases of ROS formation (25 μg/ml mitoxantrone) and ceramide abundance (25 μg/ml mitoxantrone). The effect of mitoxantrone was not significantly modified by nominal absence of extracellular Ca²⁺ but significantly blunted by the antioxidant N-acetylcysteine (1 mM). Conclusions: Mitoxantrone triggers cell membrane scrambling, an effect not requiring entry of extracellular Ca²⁺ but at least partially due to formation of ROS and ceramide.

Section: Discussionmentioning

confidence: 99%

Mitoxantrone-Induced Suicidal Erythrocyte Death

Arnold

Bissinger

Läng

2014

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“…Eryptosis may be stimulated by a wide variety of xenobiotics [15,31,32,33,34,35,36,37,38,39,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55,56,57,58,59,60,61,62,63,64,65,66] and excessive eryptosis is further observed in several clinical conditions including diabetes, renal insufficiency, hemolytic uremic syndrome, sepsis, malaria, sickle cell disease, Wilson's disease, iron deficiency, malignancy, phosphate depletion, and metabolic syndrome [15]. …”

Section: Introductionmentioning

confidence: 99%

Stimulation of Erythrocyte Cell Membrane Scrambling by Gedunin

Lupescu¹,

Bissinger²,

Warsi³

et al. 2014

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Background/Aims: Gedunin, an inhibitor of heat shock protein HSP90, triggers apoptosis of tumor cells and is thus effective against malignancy. Moreover, the drug has antimalarial potency. In analogy to apoptosis of nucleated cells, erythrocytes may enter suicidal death or eryptosis, which is characterized by cell shrinkage and by phosphatidylserine translocation to the erythrocyte surface. Eryptosis may be triggered by increase of cytosolic Ca²⁺-activity ([Ca²⁺]_i). The present study explored whether gedunin stimulates eryptosis. Methods: Forward scatter was determined to estimate cell volume, annexin V binding to identify phosphatidylserine-exposing erythrocytes, hemoglobin release to depict hemolysis, and Fluo3-fluorescence to quantify [Ca²⁺]_i. Results: A 48 h exposure of human erythrocytes to gedunin significantly increased [Ca²⁺]_i (12 µM), significantly decreased forward scatter (24 µM) and significantly increased annexin-V-binding (12 µM). The effect of gedunin (24 µM) on annexin-V-binding was virtually abrogated by removal of extracellular Ca²⁺. Conclusion: Gedunin stimulates suicidal erythrocyte death or eryptosis, an effect mainly if not exclusively due to stimulation of Ca²⁺ entry.

“…Eryptosis is triggered by a myriad of xenobiotics [26,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55,56,57,58,59,60,61,62,63,64,65,66,67,68,69,70,71,72,73,74] and excessive eryptosis is observed in a wide variety of clinical conditions, such as sepsis, malaria, sickle cell disease, Wilson's disease, iron deficiency, malignancy, metabolic syndrome, diabetes, renal insufficiency, hemolytic uremic syndrome, hyperphosphatemia and phosphate depletion [24,75]. …”

Section: Introductionmentioning

confidence: 99%

Stimulation of Eryptosis by Cryptotanshinone

Bissinger

Lupescu

Zelenak

et al. 2014

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Background/Aims: Cryptotanshinone, a component of Salvia miltiorrhiza Bunge roots, may trigger suicidal death or apoptosis of tumor cells and has thus been recommended for the prevention and treatment of malignancy. On the other hand, Cryptotanshinone has been shown to counteract apoptosis of neurons and hepatocytes. Similar to apoptosis of nucleated cells, erythrocytes may enter eryptosis, a suicidal death characterized by cell shrinkage and phosphatidylserine translocation to the erythrocyte surface. Eryptosis may be triggered by increase of cytosolic Ca²⁺-activity ([Ca²⁺]_i). The present study explored whether Cryptotanshinone stimulates eryptosis. Methods: Forward scatter was taken as measure of cell volume, annexin V binding for identification of phosphatidylserine-exposing erythrocytes and Fluo3-fluorescence for determination of [Ca²⁺]_i. Results: A 48 h exposure of human erythrocytes to Cryptotanshinone (10 µM) was followed by significant decrease of forward scatter, significant increase of the percentage annexin-V-binding cells and significant increase of [Ca²⁺]_i. The effect of Cryptotanshinone (1 µM) on annexin-V-binding was virtually abrogated by removal of extracellular Ca²⁺. Conclusion: Cryptotanshinone is a powerful stimulator of suicidal erythrocyte death or eryptosis, which is effective mainly, if not exclusively, by stimulation of Ca²⁺ entry.